Isolation and identification of bacterial strain I33M producing milk-clotting enzyme: Optimization of culture parameters using response surface

Abstract

A strain I33M which produces a milk-clotting enzyme was screened from Algerian soil near a dairy factory. This strain was identified as Bacillus mojavensis based on morphology and internal transcription spacer sequence. Sequencing analysis of 16S rDNA gene showed 100% identity of the tested strain with the B. mojavensis in the database. Phylogenetic analysis of this strain showed that it was most closely related to Bacillus subtilis strain. The optimum levels of these significant parameters to obtain the highest milk clotting activity and the lowest proteolytic activity were determined employing the response surface methodology (RSM), which revealed these as follows: wheat bran 7%, casein 0.094%, temperature 39°C, agitation size (rpm) 150. Among the various variables screened, agitation and temperature were most significant in submerged fermentation (SmF). The optimal value of milk clotting activity (MCA) is esteemed at 2.40.

Key words: Milk clotting protease, Bacillus, response surface methodology, sequencing analysis.